The phylogenetic placement of hypocrealean insect pathogens in the genus Polycephalomyces: An application of One Fungus One Name

Understanding the systematics and evolution of clavicipitoid fungi has been greatly aided by the application of molecular phylogenetics. They are now classified in three families, largely driven by reevaluation of the morphologically and ecologically diverse genus Cordyceps. Although reevaluation of morphological features of both sexual and asexual states were often found to reflect the structure of phylogenies based on molecular data, many species remain of uncertain placement due to a lack of reliable data or conflicting morphological characters. A rigid, darkly pigmented stipe and the production of a Hirsutella-like anamorph in culture were taken as evidence for the transfer of the species Cordyceps cuboidea, Cordyceps prolifica, and Cordyceps ryogamiensis to the genus Ophiocordyceps. Data from ribosomal DNA supported these species as a single group, but were unable to infer deeper relationships in Hypocreales. Here, molecular data for ribosomal and protein coding DNA from specimens of Ophiocordyceps cuboidea, Ophiocordyceps ryogamiensis, Ophiocordyceps paracuboidea, Ophiocordyceps prolifica, Cordyceps ramosopulvinata, Cordyceps nipponica, and isolates of Polycephalomyces were combined with a broadly sampled dataset of Hypocreales. Phylogenetic analyses of these data revealed that these species represent a clade distinct from the other clavicipitoid genera. Applying the recently adopted single system of nomenclature, new taxonomic combinations are proposed for these species in the genus Polycephalomyces, which has been historically reserved for asexual or anamorphic taxa.     

Phylogenetic placement of the ectomycorrhizal genus Cenococcum in Gloniaceae (Dothideomycetes)

Cenococcum is a genus of ectomycorrhizal Ascomycota that has a broad host range and geographic distribution. It is not known to produce either meiotic or mitotic spores and is known to exist only in the form of hyphae, sclerotia and host-colonized ectomycorrhizal root tips. Due to its lack of sexual and asexual spores and reproductive structures, it has proven difficult to incorporate into traditional classification within Ascomycota. Molecular phylogenetic studies of ribosomal RNA placed Cenococcum in Dothideomycetes, but the definitive identification of closely related taxa remained elusive. Here we report a phylogenetic analysis of five nuclear loci (SSU, LSU, TEF1, RPB1, RPB2) of Dothideomycetes that placed Cenococcum as a close relative of the genus Glonium of Gloniaceae (Pleosporomycetidae incertae sedis) with strong statistical support. Glonium is a genus of saprobic Dothideomycetes that produces darkly pigmented, carbonaceous, hysteriate apothecia and is not known to be biotrophic. Evolution of ectomycorhizae, Cenococcum and Dothideomycetes is discussed.     

Molecular phylogeny and new taxa in the Archaeosporales (Glomeromycota): Ambispora fennica gen. sp. nov., Ambisporaceae fam. nov., and emendation of Archaeospora and Archaeosporaceae

The AM fungal family Archaeosporaceae and the genus Archaeospora are rendered paraphyletic by the relationship with the Geosiphonaceae. This problem led to a more detailed study of the Archaeosporales. Members of the Archaeosporaceae were described as forming both glomoid and acaulosporoid spores, or solely acaulosporoid spores. However, we found that Glomus callosum fell into the same phylogenetic clade as A. leptoticha and A. gerdemannii, but exclusively formed glomoid spores. To resolve these inconsistencies, a genus, Ambispora gen. nov., typified by Ambispora fennica sp. nov., is erected based on morphological evidence and SSU and ITS region rDNA data. Ambispora contains three species known to produce both acaulosporoid and glomoid spores: A. fennica, A. leptoticha comb. nov. (basionym G. leptotichum), and A. gerdemannii comb. nov. (basionym G. gerdemannii). Another species, A. callosa comb. nov. (basionym G. callosum), is known only from glomoid spores. Ambispora is placed in a new family, the Ambisporaceae fam. nov. The Archaeosporaceae is maintained with the type species, Archaeospora trappei (basionym Acaulospora trappei), along with Intraspora schenckii (basionym Entrophospora schenckii). Acaulospora nicolsonii, known only from acaulosporoid spores, is discussed and is considered likely to belong in the Ambisporaceae, but is retained within its present genus because of inadequate morphological information and a lack of molecular data.     

Identification and Isolation of Two Ascomycete Fungi from Spores of the Arbuscular Mycorrhizal Fungus Scutellospora castanea

Two filamentous fungi with different phenotypes were isolated from crushed healthy spores or perforated dead spores of the arbuscular mycorrhizal fungus (AMF) Scutellospora castanea. Based on comparative sequence analysis of 5.8S ribosomal DNA and internal transcribed spacer fragments, one isolate, obtained from perforated dead spores only, was assigned to the genus Nectria, and the second, obtained from both healthy and dead spores, was assigned to Leptosphaeria, a genus that also contains pathogens of plants in the Brassicaceae. PCR and randomly amplified polymorphic DNA-PCR analyses, however, did not indicate similarities between pathogens and the isolate. The presence of the two isolates in both healthy spores and perforated dead spores of S. castanea was finally confirmed by transmission electron microscopy by using distinctive characteristics of the isolates and S. castanea. The role of this fungus in S. castanea spores remains unclear, but the results serve as a strong warning that sequences obtained from apparently healthy AMF spores cannot be presumed to be of glomalean origin and that this could present problems for studies on AMF genes.     

Davallia (Polypodiales: Davalliaceae) macrofossils from Early Miocene Otago (New Zealand) with in situ spores

Fossil fern fronds referable to the extant fern genus Davallia (Polypodiales: Davalliaceae) bearing sporangia with in situ spores are described from the Early Miocene Foulden Maar diatomite deposit, Otago, New Zealand. The fronds are the first published Southern Hemisphere macrofossil record for the family and provide valuable palaeoclimate data supporting warm conditions in Early Miocene New Zealand. The matching of Davallia fronds to the form spore taxon Polypodiisporites radiatus shows that the genus has had a long, apparently continuous history throughout late Cenozoic New Zealand.     

Clearance of a dermal Huffmanela sp. in a sandbar shark (Carcharhinus plumbeus) using levamisole

A wild-caught captive sandbar shark Carcharhinus plumbeus developed a contiguous network of darkly pigmented linear tracks that progressed from the snout to the ventral cervical region. Microscopic examination of a skin scraping revealed nematode eggs of the genus Huffmanela, a group of histozoic nematodes that is known to parasitize requiem sharks and marine and freshwater teleosts. The fresh eggs were darkly pigmented with bipolar plugs, contained a larva, and measured 73.3 to 86.4 by 39.0 to 47.4 microm (n = 10). Formalin-fixed and paraffin-embedded eggs were significantly smaller (Wilcoxon rank sums test, p < 0.005), measuring 70.5 to 78.9 by 33.6 to 41.3 microm (n = 13). These measurements do not correlate with previously reported species of Huffmanela. Serial treatment with levamisole (10 mg kg(-1), intramuscular [i.m.]) cleared the egg tracks within 21 d, with no recurrence or apparent complications.     

ACROSYMPHYTON FIRMUM SP. NOV. (RHODOPHYTA,CRYPTONEMIALES), A NEW SUBTIDAL RED ALGA FROM NEW ZEALAND; DEVELOPMENTAL MORPHOLOGY AND DISTRIBUTION OF THE GAMETOPHYTE1

Acrosymphyton firmum sp. nov., is described from the northeastern coast of North 1., New Zealand. Gametophytes are spring–summer annuals which grow subtidally on cobbles. Thalli are uniaxial; each axial cell bears a whorl of four indeterminate and one determinate branchlets. Indeterminate branchlets are alternately arranged giving the thallus a distichous and feather-like appearance. Numerous corticating rhizoidal filaments are produced from the periaxial and lower whorl branchlet cells. These rhizoids entwine and obscure the main axis as the thallus develops until in the mature plant the axes have a firm consistency and lubricous texture. The carpogonial branch bearing short lateral filaments and auxiliary cell branch with terminal auxiliary cell place this new species in the genus Acrosymphyton Sjöstedt. Of the three described species in the genus, A. firmum is most similar to A. taylori. This is the first report of the genus Acrosymphyton and the only confirmed report of the family Dumontiaceae in New Zealand waters.     

Ultrastructure of Minchinia Sp. Spores from Shipworms (Teredo Spp.) in the Western North Atlantic, with a Discussion of Taxonomy of the Haplosporidiidae

Electron microscopy of haplosporidan spores from Teredo navalis and T. furcifera revealed 4 distinct membrane-bound extensions, 1 apical extension opposite the opercular hinge, 1 terminal and 2 opposing lateral extensions. These extensions were not continuous with the spore wall, but contained microtubule-like structures and degrading epispore cytoplasm. No other known species in the family Haplosporidiidae is characterized by spores possessing four epispore extensions. There are currently two genera in this family, Minchinia and Haplosporidium. The genus Minchinia includes spores such as those of M. chitonis which bear two epispore cytoplasm extensions. Spores of the genus Haplosporidium have been characterized by spore wall derived filaments. A 3rd group of haplosporidan species with spores ornamented by wrappings have traditionally also been assigned to the genus Haplosporidium. Based on the presence of epispore cytoplasm extensions rather than spore wall filaments, the haplosporidan of Teredo spp. can be placed in the genus Minchinia.     

不同林龄短枝木麻黄小枝单宁含量及养分再吸收动态

通过对福建省惠安县不同林龄短枝木麻黄人工林不同发育阶段小枝单宁含量及氮磷再吸收率的研究,探讨了短枝木麻黄林分发育过程中的营养保存策略。结果表明,幼龄林(5年生)成熟小枝中的总酚(TP)、可溶性缩合单宁(ECT)总缩合单宁(TCT)含量及蛋白质结合能力(PPC)显著高于成熟林(21年生)和衰老林(38年生)。随着林分发育,成熟小枝中N含量显著升高,而P含量呈降低趋势。不同发育阶段林分成熟小枝中N:P比均大于20,且随着林龄的增加而升高。磷再吸收率(PRE)显著高于氮再吸收率(NRE),均以成熟林分最高。这表明短枝木麻黄小枝单宁含量与养分再吸收受林龄影响,其养分保存机制会随着林分生长发育的变化而进行调节。     

Lageniastrum macrosporae (fossil Volvocales, Lageniastraceae nov. fam.), an endophyte in megaspores from the Carboniferous of the French Massif Central

Sublagenicula nuda lycophyte megaspores from the upper Visean of central France frequently host a colonial green alga as an endophyte. This association was first recorded more than 100 years ago by the French paleobotanist B. Renault, who introduced the name Lageniastrum macrosporae for the alga. However, the biological significance of the discovery was not fully assessed until recently. The L. macrosporae-S. nuda association represents the oldest compelling fossil evidence for algal endophytes in land plants, and the only example to date of an alga residing in the interior of spores of vascular cryptogams. Here we present a detailed re-evaluation and photographic documentation of the surviving original specimens from the Visean of Combres/Lay and Esnost. Moreover, a newly discovered specimen from the Stephanian of central France represents the first record of this association from the Upper Carboniferous. An emended diagnosis for L. macrosporae Renault, 1896 is provided, and a lectotype and paralectotype are designated. Although L. macrosporae displays a striking suite of morphological characters found in members of the extant chlorophyte family Volvocaceae (especially the genus Volvox), the peculiar biology of the fossil necessitates establishment of a new family, for which the name Lageniastraceae is proposed. Considerations of the adaptive advantages for the alga of occupying the interior of megaspores are offered. These include the possible effectiveness of the spores in protection during periods of desiccation and against plankton-feeding animals, and use of the spores as potential vectors for dispersal from one isolated body of water to another by spore-feeding animals.     

Sexual Dichromatism of the Damselfly Calopteryx japonica Caused by a Melanin-Chitin Multilayer in the Male Wing Veins

Mature male Calopteryx japonica damselflies have dark-blue wings, due to darkly coloured wing membranes and blue reflecting veins. The membranes contain a high melanin concentration and the veins have a multilayer of melanin and chitin. Female and immature C. japonica damselflies have brown wings. We have determined the refractive index of melanin by comparing the differently pigmented wing membranes and applying Jamin-Lebedeff interference microscopy. Together with the previously measured refractive index of chitin the blue, structural colour of the male wing veins could be quantitatively explained by an optical multilayer model. The obtained melanin refractive index data will be useful in optical studies on melanized tissues, especially where melanin is concentrated in layers, thus causing iridescence.     

Phylogeny of Myxobolidae (Myxozoa) and the evolution of myxospore appendages in the Myxobolus clade

Genera Myxobolus Bütschli, 1882 and Henneguya Thélohan, 1892 (Myxobolidae) are specious myxozoan genera. They comprise nearly half of overall known myxozoan species diversity. A typical spore feature of Henneguya is the presence of two caudal appendages of the spore valves, which distinguishes them from species of the genus Myxobolus. Several Myxobolus spp., however, were reported to show aberrant spores with Henneguya-like caudal appendages. We found such aberrant spores in Myxobolus tsangwuensis and Myxobolus wulii. We studied the ultrastructure of M. wulii and Myxobolus oralis spores with caudal appendages by transmission electron microscopy (TEM). TEM of these aberrant spores revealed that their caudal appendages have the same ultrastructure as the appendages of Henneguya spp. Small caudal appendages of M. wulii spores observed only on TEM suggested that this character may be often overlooked and more Myxobolus species potentially have the ability to express the caudal appendages on the myxospore. In order to trace the evolution of this character, we performed broad phylogenetic analysis of all species of the family Myxobolidae which are available in GenBank including nearly 300 taxa. We found at least eight independent evolutionary origins of spores with two appendages, three origins of a single appendage and 12 apparent secondary losses of the spore projections. Therefore, genus Henneguya with typical two-tailed myxospores is polyphyletic, however a majority of its species has a common ancestor and groups in the second largest subclade of the Myxobolus clade. We also mapped the biological characteristics (host, site of infection and environment) of Myxobolidae species on the phylogenetic tree. We revealed an evident host-associated evolutionary pattern in all parts of the Myxobolus clade with a distinct and species-rich subclade containing almost exclusively species infecting species of the Order Cypriniformes.     

On the systematic position of the Early Cretaceous fern genus “Athyrium”

The oldest fossils assigned to Athyrium (mostly based on the sorus morphology) comprise fronds and spores from the Lower Cretaceous of Northeast Asia. However, most molecular dating suggests that extant Athyrium diverged from its sister genus during the Eocene or later, implying that the Cretaceous fossils probably belong to another polypodiaceous taxon. By examining the sorus morphology of extant genera related to the family Athyriaceae, we found that the primary diagnostic feature for assigning the Cretaceous fossils to Athyrium, i.e., the sorus shape, is common to the entire extant family, or plesiomorphic for the genus. As the fronds are more commonly preserved than the reproductive parts, we compared the fossil frond morphology with those of living taxa of the family that is divided into two types. The Cretaceous fossil we examined here bears the frond’s costal groove characters on adaxial side, which is more closely related to that of the Deparia-clade instead of the clade including Athyrium and other genera of the family. The observation is further confirmed by the cladistic analysis using morphological characters. The systematic position of the Early Cretaceous “Athyrium” was resolved as a stem member of the total Athyriaceae using a tip-dating approach with the Fossilized Birth-Death model in a Bayesian framework. Our study suggests that Early Cretaceous fossils previously assigned to Athyrium require taxonomic revision.     

Spore morphology of the genus Dryopteris Adans. (Dryopteridaceae) in Korea

Spore morphology of the Korean members of the genus Dryopteris was examined by scanning electron microscopy. In particular, spores of D. hangchowensis, D. woodsiisora, D. saxifragivaria, and D. subexaltata were examined here for the first time. Three perispore types were recognized on the basis of the shape of macro-ornamentations on perispore: these include rugate, echinate, and spinose. The rugate perispore type appeared to be the most common in the genus, and probably represents plesiomorphic condition in the genus Dryopteris. The echinate perispore type is found in D. expansa, and spinose perispore type in D. formosana. The results of this study demonstrate that the perispore characteristics, particularly the shape of macro-ornamentation and surface texture, have diagnostic value at the species level. Indeed, most of the Dryopteris taxa in Korea could be distinguished by their perispore characters. In addition, spores of sexual diploid individuals of D. saxifraga were much smaller than those of agamosporous triploid individuals, supporting the contention that the size of spores in ferns is generally related to ploidy levels.     

Morphology and ultrasctructure of spores Klukia tyganensis Krassilov (Schizaeaceae, Filicales) from the Berriassian of the Tyrma Depression (Russian Far East)

In situ spores of the fern Klukia tyganensis Krassil. from the coal-bearing deposits on the right bank of the Tyrma River (Berriassian, Bureya River Basin, Tyrma Depression) have been examined using light, scanning and transmission electron microscopy. The spores of K. tyganensis are demonstrated to be similar in the ultrastructure of sporoderm to that of some species of the extant genus Anemia and to differ from the spores of Lygodium in the ultrastructure of perispore and exospore.     

Biflavones of Dacrydium sensu lato

A survey of the biflavonyls in the leaves and branchlets of all three sections of the genus Dacrydium s.l. revealed a complex mixture of amentoflavone and its partial methyl ethers and hinokiflavone in most species. A group of three species in section C, however, were characterised by cupressuflavone derivatives as the major biflavone constituents. The results are used to discuss proposals for revised generic boundaries.     

Heterokaryosis and the role of cytoplasmic inheritance in dark resting structure formation in Verticillium spp

Summary Auxotrophic and morphological mutants of Verticillium albo-atrum (producing darkly pigmented resting mycelium) and V. dahliae (forming dark microsclerotia) were isolated after treatment of conidia (haploid and uninucleate) with ultraviolet light. Hyphal tip and conidial analysis revealed that complementation between pairs of auxotrophs on minimal medium was due to a mosaic of homokaryotic and heterokaryotic regions with some hyphal tips growing syntrophically. A degree of incompatibility was observed in a few intraspecific, but in most of the interspecific, heterokaryon tests. Heterozygous diploid conidia (6–11 in length compared with 3–6 for haploids) were recovered at a frequency of 1 in 8x10⁶ by plating spores at high density on MM. Young diploid colonies segregated to give haploid and diploid sectors, some of which were recombinant types (parasexual cycle). Heterokaryons between complementary auxotrophs which were wild-type for dark pigmentation (hyl⁺) resembled wild-type and only darkly pigmented colonies were recovered by conidial analysis. Heterokaryons between hyl⁺ and hyaline (hyl) auxotrophs again resembled hyl⁺ morphology and usually only hyl⁺ colonies of both auxotrophic genotypes were recovered. Conidia from heterokaryons formed by stable hyl auxotrophs produced only hyl colonies of both auxotrophic genotypes. The important role played by cytoplasmic factors in the inheritance of darkly-pigmented resting structures in Verticillium was strongly confirmed by the present work.     

Two new species of Kladnostrobus nov. gen. and their spores from the Pennsylvanian of the Kladno-Rakovník Basin (Bolsovian, Czech Republic)

A new lycopsid family Kladnostrobaceae is proposed, based on the type of sporangia, their attachment by a pedicel and the type of reticulate spores enclosed. All these characteristics distinguish the Kladnostrobaceae from all other lycopsid families. A new lycopsid genus Kladnostrobus nov. gen., consisting of two new species Kladnostrobus clealii nov. sp. and Kladnostrobus psendae nov. sp., is described from the Kladno-Rakovník Basin (Lower Bolsovian) of the central and western Carboniferous continental basins of the Czech Republic. Helically arranged distal laminae and pedicels are relatively primitive, suggesting that Kladnostrobus may represent a new, primitive type of lycopsid cone produced by some unknown, probably arborescent lycopsid parent plant. Spores of Kladnostrobus are about 90-100 μm in diameter, and possess reticulate sculpture. The proximal contact area of spores is laevigate. In situ spores can resemble some dispersed species of the genera Convolutispora Hoffmeister, Staplin and Malloy, Camptotriletes (Naumova) Potonié and Kremp, Reticulatisporites (Ibrahim) Neves and mainly Dictyotriletes (Naumova) Smith and Buttterworth.